GPCRs are one of the largest gene families in the human genome and have been the most tractable set of targets for the development of clinically effective, small molecule, medicines. It is estimated that of the drugs used clinically in man some 40% target GPCRs. There is thus great interest in the details of the structure, regulation and activation mechanisms of GPCRs as well as the downstream signalling cascades they control. The class A or rhodopsin-like family of GPCRs is by far the largest containing more than 80% of the total GPCR family members. More than 800 genes encoding GPCRs have been identified in the human genome sequencing programme but only some 25 of these are currently the target for clinically effective medicines. There is thus great potential to expand this and to find useful medicines that target recently identified GPCRs (Lee et al., 2001).
In the recent past, the concept that GPCRs exist as dimers has moved rapidly from hypothesis to clearly accepted (see Bouvier, 2001, Milligan, 2001, George et al., 2002 for reviews). Although homodimers (i.e. a dimer containing two copies of one individual GPCR) have been the best studied, growing evidence suggests that heterodimerisation (i.e. the dimer consists of one molecule of each of two different GPCRs) both occurs and can have both functional and pharmacological sequelae (Devi, 2001, George et al., 2002). However, important questions remain in relation to the selectivity of formation of such heterodimers and how to monitor the function of a heterodimer in isolation when co-expression of two different GPCRs must also result in the production of homodimeric pairs. Given that many GPCRs are co-expressed in a single cell then it is likely that the complement of GPCR dimers in a cell is complex.
Studies have been carried out on the γ-aminobutyric acid (GABA) type B receptor (GABABR) (Duthey et al., 2002). This is an unusual GPCR because it is the only one known to date that needs two subunits, GB1 and GB2, to function. The GB1 subunit contains the GABA binding site but is unable to activate G-protein alone. GB2 does not bind GABA but does have the ability to activate G-proteins. Duthey et al. looked at the role of each subunit within the GB1-GB2 heteromer in G-protein coupling. The study included introducing mutations into both GB1 and GB2, particularly within the third intracellular loop. They determined that mutation to GB2 prevents G-protein activation, whereas a similar mutation to GB1 did not affect receptor function. Although interesting for the GABAB receptor, this study unfortunately does not provide any information on GPCRs where the same protein is responsible for both ligand binding and G-protein activation.
Further studies looked at the co-expression of a first mutant receptor which was defective in hormone binding and a second mutant receptor which was defective in signal generation. It was reported that co-expression of the two mutants rescued hormone-activated cAMP production (Lee et al., J. Biol. Chem. Vol. 277, No. 18, 2002; Osuga et al., J. Biol. Chem. Vol. 272, No. 40, 1997).
However, although it is acknowledged that GPCRs are extremely important as potential drug targets, there does not exist a satisfactory screening assay which allows reliable data to be gathered about the functional properties, e.g. ligand binding properties, of potentially naturally occurring GPCR oligomers, particularly GPCR hetero-oligomers.